Intracellular Streptococcus pyogenes in Human Macrophages Display an Altered Gene Expression Profile
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{"title"=>"Intracellular Streptococcus pyogenes in Human Macrophages Display an Altered Gene Expression Profile", "type"=>"journal", "authors"=>[{"first_name"=>"Erika", "last_name"=>"Hertzén"}, {"first_name"=>"Linda", "last_name"=>"Johansson"}, {"first_name"=>"Rita", "last_name"=>"Kansal"}, {"first_name"=>"Alexander", "last_name"=>"Hecht"}, {"first_name"=>"Samira", "last_name"=>"Dahesh"}, {"first_name"=>"Marton", "last_name"=>"Janos"}, {"first_name"=>"Victor", "last_name"=>"Nizet"}, {"first_name"=>"Malak", "last_name"=>"Kotb"}, {"first_name"=>"Anna", "last_name"=>"Norrby-Teglund"}], "year"=>2012, "source"=>"PLoS ONE", "identifiers"=>{"scopus"=>"2-s2.0-84871898477", "doi"=>"10.1371/journal.pone.0035218", "issn"=>"1932-6203", "pui"=>"366360196", "sgr"=>"84871898477", "pmid"=>"22511985"}, "id"=>"50cf0404-ce7b-3510-904c-a5469da317bb", "abstract"=>"Streptococcus pyogenes is an important human pathogen, which has recently gained recognition as an intracellular microorganism during the course of severe invasive infections such as necrotizing fasciitis. Although the surface anchored M protein has been identified as a pivotal factor affecting phagosomal maturation and S. pyogenes survival within macrophages, the overall transcriptional profile required for the pathogen to adapt and persist intracellularly is as of yet unknown. To address this, the gene expression profile of S. pyogenes within human macrophages was determined and compared to that of extracellular bacteria using customized microarrays and real-time qRT-PCR. In order to model the early phase of infection involving adaptation to the intracellular compartment, samples were collected 2h post-infection. Microarray analysis revealed that the expression of 145 streptococcal genes was significantly altered in the intracellular environment. The majority of differentially regulated genes were associated with metabolic and energy-dependent processes. Key up-regulated genes in early phase intracellular bacteria were ihk and irr, encoding a two-component gene regulatory system (TCS). Comparison of gene expression of selected genes at 2h and 6h post-infection revealed a dramatic shift in response regulators over time with a down-regulation of ihk/irr genes concurring with an up-regulation of the covR/S TCS. In re-infection assays, intracellular bacteria from the 6h time point exhibited significantly greater survival within macrophages than did bacteria collected at the 2h time point. An isogenic S. pyogenes mutant deficient in ihk/irr displayed significantly reduced bacterial counts when compared to wild-type bacteria following infection of macrophages. The findings illustrate how gene expression of S. pyogenes during the intracellular life cycle is fine-tuned by temporal expression of specific two-component systems.; ", "link"=>"http://www.mendeley.com/research/intracellular-streptococcus-pyogenes-human-macrophages-display-altered-gene-expression-profile", "reader_count"=>8, "reader_count_by_academic_status"=>{"Researcher"=>2, "Student > Ph. D. Student"=>3, "Student > Postgraduate"=>2, "Student > Bachelor"=>1}, "reader_count_by_user_role"=>{"Researcher"=>2, "Student > Ph. D. Student"=>3, "Student > Postgraduate"=>2, "Student > Bachelor"=>1}, "reader_count_by_subject_area"=>{"Agricultural and Biological Sciences"=>3, "Medicine and Dentistry"=>2, "Psychology"=>1, "Immunology and Microbiology"=>2}, "reader_count_by_subdiscipline"=>{"Medicine and Dentistry"=>{"Medicine and Dentistry"=>2}, "Psychology"=>{"Psychology"=>1}, "Immunology and Microbiology"=>{"Immunology and Microbiology"=>2}, "Agricultural and Biological Sciences"=>{"Agricultural and Biological Sciences"=>3}}, "reader_count_by_country"=>{"United States"=>1}, "group_count"=>0}

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Figshare

  • {"files"=>["https://ndownloader.figshare.com/files/654300"], "description"=>"<p>Monocyte-derived macrophages were infected for various time points with either 5448 WT strain or an isogenic deletion mutant (Δihk/irr). The cells were infected with equal infectious doses and the infection was allowed to proceed for 1-2h depending on assay after which extracellular bacteria were eradicated by antibiotics. <b>A</b>) Colony forming units plated from macrophage lysates at 2h pi according to <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035218#s4\" target=\"_blank\">material and methods</a>. Data from 7 donors, <b>B</b>) shows the mean % ± SEM of cells harboring viable cocci as determined by bacterial viability stain at 4 h pi (n = 7 donors), <b>C</b>) Percent dead cell marker positive cells at various time points after infection as analyzed by flow cytometry (n = 2 donors). The bars show mean ± SEM of duplicate samples from one representative donor. <b>D</b>) At 4h pi, antibiotic-free media was added and bacterial load as monitored by plating cell culture media at defined time points shows the results of infection of cells from one representative donor, and (<b>E</b>) compiled data from 5 experiments using cells from different donors (indicated by separate symbols). Note that due to the log scale, some symbols overlap. Statistically significant differences were determined by use of paired Wilcoxon signed rank test and <i>p</i> values are indicated in the graphs.</p>", "links"=>[], "tags"=>["bacterial", "counts", "compared", "wildtype"], "article_id"=>324777, "categories"=>["Microbiology", "Biotechnology", "Genetics", "Infectious Diseases"], "users"=>["Erika Hertzén", "Linda Johansson", "Rita Kansal", "Alexander Hecht", "Samira Dahesh", "Marton Janos", "Victor Nizet", "Malak Kotb", "Anna Norrby-Teglund"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0035218.g004", "stats"=>{"downloads"=>0, "page_views"=>7, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Reduced_bacterial_counts_of_an_ihk_irr_deficient_mutant_as_compared_to_wildtype_S_pyogenes_/324777", "title"=>"Reduced bacterial counts of an ihk/irr-deficient mutant, as compared to wildtype <b>S. pyogenes.</b>", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-04-12 01:19:37"}
  • {"files"=>["https://ndownloader.figshare.com/files/654477"], "description"=>"a<p>Microarray array data.</p>b<p>Gene expression in wildtype and <i>irr</i>-deficient <i>S. pyogenes</i> strains was determined at both early and late exponential growth phase, indicated by EE and LE resp.</p><p>Genes that show the same regulatory pattern in this study and in Voyich et al. <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035218#pone.0035218-Voyich2\" target=\"_blank\">[25]</a> are indicated in bold.</p>", "links"=>[], "tags"=>["regulated", "genes", "differentially", "intracellular", "wildtype", "2h"], "article_id"=>324954, "categories"=>["Microbiology", "Biotechnology", "Genetics", "Infectious Diseases"], "users"=>["Erika Hertzén", "Linda Johansson", "Rita Kansal", "Alexander Hecht", "Samira Dahesh", "Marton Janos", "Victor Nizet", "Malak Kotb", "Anna Norrby-Teglund"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0035218.t001", "stats"=>{"downloads"=>1, "page_views"=>8, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Comparison_of_previously_identified_Ihk_Irr_regulated_genes_25_with_differentially_expressed_genes_of_intracellular_wildtype_bacteria_at_2h_pi_/324954", "title"=>"Comparison of previously identified Ihk/Irr regulated genes [25] with differentially expressed genes of intracellular wildtype bacteria at 2h pi.", "pos_in_sequence"=>0, "defined_type"=>3, "published_date"=>"2012-04-12 01:22:34"}
  • {"files"=>["https://ndownloader.figshare.com/files/654070"], "description"=>"<p><b>A</b>) Expression of <i>ihk, irr,</i> and ABC transporters in intracellular 2h pi and extracellular bacteria determined by qRT-PCR analysis. Normalized mean value of 4 biological replicates shown as fold change in intra- versus extracellular bacteria. <b>B</b>) Expression of selected genes at 2h pi determined by qRT-PCR or microarray. RNA from 3 - 4 biological replicates per method was analyzed. The normalized mean value for each gene is found in the table.</p>", "links"=>[], "tags"=>["transporter", "up-regulation", "confirmed"], "article_id"=>324552, "categories"=>["Microbiology", "Biotechnology", "Genetics", "Infectious Diseases"], "users"=>["Erika Hertzén", "Linda Johansson", "Rita Kansal", "Alexander Hecht", "Samira Dahesh", "Marton Janos", "Victor Nizet", "Malak Kotb", "Anna Norrby-Teglund"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0035218.g002", "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Two_component_system_and_transporter_up_regulation_confirmed_with_qRT_PCR_/324552", "title"=>"Two component system and transporter up-regulation confirmed with qRT-PCR.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-04-12 01:15:52"}
  • {"files"=>["https://ndownloader.figshare.com/files/653984"], "description"=>"<p><b>A</b>) Heat map of genes (n = 145) that displayed significantly (<i>P</i> : < 0.05) altered expression in intracellular versus extracellular <i>S. pyogenes</i> at 2h pi. RNA from 3 biological replicates were hybridized to microarrays and analyzed as described in the <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035218#s4\" target=\"_blank\">material and method</a> section. Red color indicates up-regulation of genes; green down-regulation and black colored genes are not active as compared to house-keeping genes in the respective condition. <b>B</b>) Bar graph displaying differently expressed genes according to functional categories. Red bars indicate ≥75% of the genes are up-regulated, green bars indicate that ≥75% of the genes are down-regulated and black bars indicate that there is no consistent trend within the category.</p>", "links"=>[], "tags"=>["intracellular", "extracellular"], "article_id"=>324465, "categories"=>["Microbiology", "Biotechnology", "Genetics", "Infectious Diseases"], "users"=>["Erika Hertzén", "Linda Johansson", "Rita Kansal", "Alexander Hecht", "Samira Dahesh", "Marton Janos", "Victor Nizet", "Malak Kotb", "Anna Norrby-Teglund"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0035218.g001", "stats"=>{"downloads"=>0, "page_views"=>6, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Differential_gene_expression_profile_in_intracellular_versus_extracellular_bacteria_/324465", "title"=>"Differential gene expression profile in intracellular versus extracellular bacteria.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-04-12 01:14:25"}
  • {"files"=>["https://ndownloader.figshare.com/files/654152"], "description"=>"<p>RNA from intracellular and extracellular <i>S. pyogenes</i> was isolated at 2h and 6h pi. <b>A</b>) shows qRT-PCR data from <i>ihk/irr, </i><b>B</b>) <i>covR/S,</i> and <b>C</b>) <i>emm1.</i> 6 biological replicates were analyzed with each sample in triplicate per gene. Statistical significant differences were determined by Wilcoxon signed rank test, and <i>p</i> values are indicated in the figure. Additional experiments included one additional time point at 1h pi and was analyzed for two donors showing <i>ihk/irr </i><b>D</b>), <i>covR/S </i><b>E</b>) and <i>emm1 </i><b>F</b>). Monocyte-derived macrophages were infected for 2h and 6h, respectively, after which the cells were lysed and the lysate added to new macrophages. Two hours post infection the cells were lysed and intracellular bacterial counts determined. <b>G</b>) shows a schematic illustration of this reinfection as well as data on intracellular bacteria 2h post infection as determined by plating of cell lysates.</p>", "links"=>[], "tags"=>["systems", "proceeds"], "article_id"=>324636, "categories"=>["Microbiology", "Biotechnology", "Genetics", "Infectious Diseases"], "users"=>["Erika Hertzén", "Linda Johansson", "Rita Kansal", "Alexander Hecht", "Samira Dahesh", "Marton Janos", "Victor Nizet", "Malak Kotb", "Anna Norrby-Teglund"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0035218.g003", "stats"=>{"downloads"=>0, "page_views"=>6, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Shift_in_gene_regulatory_systems_as_the_infection_proceeds_influence_infectivity_/324636", "title"=>"Shift in gene regulatory systems as the infection proceeds influence infectivity.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-04-12 01:17:16"}
  • {"files"=>["https://ndownloader.figshare.com/files/336035"], "description"=>"<div><p><em>Streptococcus pyogenes</em> is an important human pathogen, which has recently gained recognition as an intracellular microorganism during the course of severe invasive infections such as necrotizing fasciitis. Although the surface anchored M protein has been identified as a pivotal factor affecting phagosomal maturation and <em>S. pyogenes</em> survival within macrophages, the overall transcriptional profile required for the pathogen to adapt and persist intracellularly is as of yet unknown. To address this, the gene expression profile of <em>S. pyogenes</em> within human macrophages was determined and compared to that of extracellular bacteria using customized microarrays and real-time qRT-PCR. In order to model the early phase of infection involving adaptation to the intracellular compartment, samples were collected 2h post-infection. Microarray analysis revealed that the expression of 145 streptococcal genes was significantly altered in the intracellular environment. The majority of differentially regulated genes were associated with metabolic and energy-dependent processes. Key up-regulated genes in early phase intracellular bacteria were <em>ihk</em> and <em>irr</em>, encoding a two-component gene regulatory system (TCS). Comparison of gene expression of selected genes at 2h and 6h post-infection revealed a dramatic shift in response regulators over time with a down-regulation of <em>ihk/irr</em> genes concurring with an up-regulation of the <em>cov</em>R/S TCS. In re-infection assays, intracellular bacteria from the 6h time point exhibited significantly greater survival within macrophages than did bacteria collected at the 2h time point. An isogenic <em>S. pyogenes</em> mutant deficient in <em>ihk/irr</em> displayed significantly reduced bacterial counts when compared to wild-type bacteria following infection of macrophages. The findings illustrate how gene expression of <em>S. pyogenes</em> during the intracellular life cycle is fine-tuned by temporal expression of specific two-component systems.</p> </div>", "links"=>[], "tags"=>["intracellular", "macrophages", "altered"], "article_id"=>126396, "categories"=>["Biotechnology", "Cancer", "Genetics", "Microbiology"], "users"=>["Erika Hertzén", "Linda Johansson", "Rita Kansal", "Alexander Hecht", "Samira Dahesh", "Marton Janos", "Victor Nizet", "Malak Kotb", "Anna Norrby-Teglund"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0035218", "stats"=>{"downloads"=>1, "page_views"=>16, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/Intracellular_Streptococcus_pyogenes_in_Human_Macrophages_Display_an_Altered_Gene_Expression_Profile/126396", "title"=>"Intracellular <em>Streptococcus pyogenes</em> in Human Macrophages Display an Altered Gene Expression Profile", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-04-12 01:46:36"}

PMC Usage Stats | Further Information

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Relative Metric

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